This application is conceived on the premise that in certain diseases, e.g. peripheral artery disease (PAD), traditional therapies are ineffective in the treatment of symptoms and related morbidities. PAD currently impacts over 8 million Americans and will affect many more as our nation ages. The health care costs to treat subjects are significant ($4 billion annually by Medicare), with most patients requiring numerous endovascular and surgical procedures including amputation. Medical therapy has failed to impact the trend in this cost and efficacy curve. This proposal seeks to change this paradigm by recognizing the endothelial dysfunction attributed to PAD prevents proper blood flow recovery because the production of nitric oxide through its cognate enzyme nitric oxide synthase is impaired. We have found nitric oxide (NO) precursors formed within endothelial vesicles to be stimulated by red light energy to release NO and produce a stable NO bound vasodilator. Most importantly the production and release of this autocrine factor dilates arteries/arterioles in the absence of nitric oxide synthase (NOS). The actions of red light are clinically relevant, as we have identified significant elevations in blood flow when the gastrocnemius muscle of healthy subjects and patients with PAD are exposed to red light. The approach in this application will be to assess the production of these thiol based NO vesicles in healthy conditions and oxidative stress by using ex vivo and cell culture techniques, characterize the intracellular mechanism for vesicle transport and release of the NO precursor vesicle, and confirm our findings in vessels obtained from human subjects with and without diseases predisposed to oxidative stress. We expect the data collected will identify the mechanisms by which vesicles containing NO precursor molecules can be increased, improve endothelial dysfunction and enhance perfusion. RELEVANCE TO VETERANS: PAD is a costly and highly morbid disease which disproportionately affects our veterans. Red light has the potential to noninvasively increase blood flow through the manipulation of intracellular NO precursors produced in vesicles. This finding is a novel intervention which has great potential to reduce costs and improve symptoms by avoiding morbid surgical procedures. The broader clinical impact of red light therapy cannot be fully realized until the mechanism by which red light releases this vasodilator mechanism is identified.